title: G protein modulation of K2P potassium channel TASK-2 : a role of basic residues in the C terminus domain.
abstract: TASK-2 (K2P5.1) is a background K(+) channel opened by extra- or intracellular alkalinisation that plays a role in renal bicarbonate handling, central chemoreception and cell volume regulation. Here, we present results that suggest that TASK-2 is also modulated by Gβγ subunits of heterotrimeric G protein. TASK-2 was strongly inhibited when GTP-γ-S was used as a replacement for intracellular GTP. No inhibition was present using GDP-β-S instead. Purified Gβγ introduced intracellularly also inhibited TASK-2 independently of whether GTP or GDP-β-S was present. The effects of GTP-γ-S and Gβγ subunits were abolished by neutralisation of TASK-2 C terminus double lysine residues K257-K258 or K296-K297. Use of membrane yeast two hybrid (MYTH) experiments and immunoprecipitation assays using tagged proteins gave evidence for a physical interaction between Gβ1 and Gβ2 subunits and TASK-2, in agreement with expression of these subunits in proximal tubule cells. Co-immunoprecipitation was impeded by mutating C terminus K257-K258 (but not K296-K297) to alanines. Gating by extra- or intracellular pH was unaltered in GTP-γ-S-insensitive TASK-2-K257A-K258A mutant. Shrinking TASK-2-expressing cells in hypertonic solution decreased the current to 36 % of its initial value. The same manoeuvre had a significantly diminished effect on TASK-2-K257A-K258A- or TASK-2-K296-K297-expressing cells, or in cells containing intracellular GDP-β-S. Our data are compatible with the concept that TASK-2 channels are modulated by Gβγ subunits of heterotrimeric G protein. We propose that this modulation is a novel way in which TASK-2 can be tuned to its physiological functions.
authors.affiliation: Centro de Estudios Científicos (CECs), Avenida Arturo Prat 514, Valdivia, Chile.
authors.forename: Carolina
authors.initials: C
authors.lastname: Añazco
authors.forename: Gaspar
authors.initials: G
authors.lastname: Peña-Münzenmayer
authors.forename: Carla
authors.initials: C
authors.lastname: Araya
authors.forename: L Pablo
authors.initials: LP
authors.lastname: Cid
authors.forename: Francisco V
authors.initials: FV
authors.lastname: Sepúlveda
authors.forename: María Isabel
authors.initials: MI
authors.lastname: Niemeyer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D044387:GTP-Binding Protein beta Subunits / Q000235:genetics / Q000378:metabolism*; D044388:GTP-Binding Protein gamma Subunits / Q000235:genetics / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D057809:HEK293 Cells; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D051379:Mice; D024683:Potassium Channels, Tandem Pore Domain / Q000378:metabolism*; D013873:Thionucleotides / Q000494:pharmacology; D020798:Two-Hybrid System Techniques
chemical_list: D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; C109137:KCNK1 protein, human; C498739:Kcnk5 protein, mouse; D024683:Potassium Channels, Tandem Pore Domain; D013873:Thionucleotides; D006153:Guanosine Diphosphate; C023427:guanosine 5'-O-(2-thiodiphosphate); D020962:Heterotrimeric GTP-Binding Proteins